Stephen P. Mulligan, M.B., B.S., Ph.D. University of Sydney (Australia)

IMMUNOLOGY

Global phenotype profiling of chronic lymphocytic leukemia

Update:

This project headed by myself and colleagues at the University of Sydney attempts to give additional information on the proteins which are expressed on CLL cells. The proteins on the surface of the cell identify whether they are T-lymphocytes or B-lymphocytes and also whether they are in an activated or an inactive form. The traditional methodology of trying to establish the protein profile on the cells is by conventional flow cytometry. We developed a machine that characterizes the proteins on the cells' surface. The cells are incubated with fluorescent antibodies which after going through this machine will send a signal if the protein is on the surface.

Antibodies used in this flow cytometry technique are expensive and there is a limit to the number of samples that can be done at any one time. This project is analyzing 147 proteins. These proteins were selected based on some exploratory work which was conducted by our group. 50 CLL patients have been explored and we have begun a large scale study of patients with known treatment and survival outcomes.

ZAP-70 is an important protein which switches on CLL cells. The methodology to accurately access the amount of ZAP-70 and its activity in CLL cells is complicated and there is a lack of standardization. Our group has developed a method for identifying ZAP-70 by measuring the amount in cell extracts. Linking ZAP-70 with all the other surface proteins on the cells will give more accurate information regarding the level of activity, whether the cells are likely to be susceptible to different treatments and overall outcome. This method is reproducible and considerably cheaper than any other methods which have been developed to identify the protein profile of CLL cells.